The exception that proves the rule: Virulence gene expression at the onset of Plasmodium falciparum blood stage infections.

Controlled human malaria infections (CHMI) are a valuable tool to study parasite gene expression in vivo under defined conditions. In previous studies, virulence gene expression was analyzed in samples from volunteers infected with the Plasmodium falciparum (Pf) NF54 isolate, which is of African origin. Here, we provide an in-depth investigation of parasite virulence gene expression in malaria-naïve European volunteers undergoing CHMI with the genetically distinct Pf 7G8 clone, originating in Brazil. Differential expression of var genes, encoding major virulence factors of Pf, PfEMP1s, was assessed in ex vivo parasite samples as well as in parasites from the in vitro cell bank culture that was used to generate the sporozoites (SPZ) for CHMI (Sanaria PfSPZ Challenge (7G8)). We report broad activation of mainly B-type subtelomeric located var genes at the onset of a 7G8 blood stage infection in naïve volunteers, mirroring the NF54 expression study and suggesting that the expression of virulence-associated genes is generally reset during transmission from the mosquito to the human host. However, in 7G8 parasites, we additionally detected a continuously expressed single C-type variant, Pf7G8_040025600, that was most highly expressed in both pre-mosquito cell bank and volunteer samples, suggesting that 7G8, unlike NF54, maintains expression of some previously expressed var variants during transmission. This suggests that in a new host, the parasite may preferentially express the variants that previously allowed successful infection and transmission. Trial registration: ClinicalTrials.gov - NCT02704533; 2018-004523-36.

Sterile protection against human malaria by chemoattenuated PfSPZ vaccine.

A highly protective malaria vaccine would greatly facilitate the prevention and elimination of malaria and containment of drug-resistant parasites. A high level (more than 90%) of protection against malaria in humans has previously been achieved only by immunization with radiation-attenuated Plasmodium falciparum (Pf) sporozoites (PfSPZ) inoculated by mosquitoes; by intravenous injection of aseptic, purified, radiation-attenuated, cryopreserved PfSPZ ('PfSPZ Vaccine'); or by infectious PfSPZ inoculated by mosquitoes to volunteers taking chloroquine or mefloquine (chemoprophylaxis with sporozoites). We assessed immunization by direct venous inoculation of aseptic, purified, cryopreserved, non-irradiated PfSPZ ('PfSPZ Challenge') to malaria-naive, healthy adult volunteers taking chloroquine for antimalarial chemoprophylaxis (vaccine approach denoted as PfSPZ-CVac). Three doses of 5.12 × 104 PfSPZ of PfSPZ Challenge at 28-day intervals were well tolerated and safe, and prevented infection in 9 out of 9 (100%) volunteers who underwent controlled human malaria infection ten weeks after the last dose (group III). Protective efficacy was dependent on dose and regimen. Immunization with 3.2 × 103 (group I) or 1.28 × 104 (group II) PfSPZ protected 3 out of 9 (33%) or 6 out of 9 (67%) volunteers, respectively. Three doses of 5.12 × 104 PfSPZ at five-day intervals protected 5 out of 8 (63%) volunteers. The frequency of Pf-specific polyfunctional CD4 memory T cells was associated with protection. On a 7,455 peptide Pf proteome array, immune sera from at least 5 out of 9 group III vaccinees recognized each of 22 proteins. PfSPZ-CVac is a highly efficacious vaccine candidate; when we are able to optimize the immunization regimen (dose, interval between doses, and drug partner), this vaccine could be used for combination mass drug administration and a mass vaccination program approach to eliminate malaria from geographically defined areas.

The Adjuvanted Recombinant Zoster Vaccine Confers Long-Term Protection Against Herpes Zoster: Interim Results of an Extension Study of the Pivotal Phase 3 Clinical Trials ZOE-50 and ZOE-70.

BACKGROUND: This ongoing follow-up study evaluated the persistence of efficacy and immune responses for 6 additional years in adults vaccinated with the glycoprotein E (gE)-based adjuvanted recombinant zoster vaccine (RZV) at age ≥50 years in 2 pivotal efficacy trials (ZOE-50 and ZOE-70). The present interim analysis was performed after ≥2 additional years of follow-up (between 5.1 and 7.1 years [mean] post-vaccination) and includes partial data for year (Y) 8 post-vaccination. METHODS: Annual assessments were performed for efficacy against herpes zoster (HZ) from Y6 post-vaccination and for anti-gE antibody concentrations and gE-specific CD4[2+] T-cell (expressing ≥2 of 4 assessed activation markers) frequencies from Y5 post-vaccination. RESULTS: Of 7413 participants enrolled for the long-term efficacy assessment, 7277 (mean age at vaccination, 67.2 years), 813, and 108 were included in the cohorts evaluating efficacy, humoral immune responses, and cell-mediated immune responses, respectively. Efficacy of RZV against HZ through this interim analysis was 84.0% (95% confidence interval [CI], 75.9-89.8) from the start of this follow-up study and 90.9% (95% CI, 88.2-93.2) from vaccination in ZOE-50/70. Annual vaccine efficacy estimates were >84% for each year since vaccination and remained stable through this interim analysis. Anti-gE antibody geometric mean concentrations and median frequencies of gE-specific CD4[2+] T cells reached a plateau at approximately 6-fold above pre-vaccination levels. CONCLUSIONS: Efficacy against HZ and immune responses to RZV remained high, suggesting that the clinical benefit of RZV in older adults is sustained for at least 7 years post-vaccination. Clinical Trials Registration. NCT02723773.

Cellular and antibody response in GMZ2-vaccinated Gabonese volunteers in a controlled human malaria infection trial.

BACKGROUND: Antibody and cellular memory responses following vaccination are important measures of immunogenicity. These immune markers were quantified in the framework of a vaccine trial investigating the malaria vaccine candidate GMZ2. METHODS: Fifty Gabonese adults were vaccinated with two formulations (aluminum Alhydrogel and CAF01) of GMZ2 or a control vaccine (Verorab). Vaccine efficacy was assessed using controlled human malaria infection (CHMI) by direct venous inoculation of 3200 live Plasmodium falciparum sporozoites (PfSPZ Challenge). GMZ2-stimulated T and specific B-cell responses were estimated by flow cytometry before and after vaccination. Additionally, the antibody response against 212 P. falciparum antigens was estimated before CHMI by protein microarray. RESULTS: Frequencies of pro- and anti-inflammatory CD4+ T cells stimulated with the vaccine antigen GMZ2 as well as B cell profiles did not change after vaccination. IL-10-producing CD4+ T cells and CD20+ IgG+ B cells were increased post-vaccination regardless of the intervention, thus could not be specifically attributed to any malaria vaccine regimen. In contrast, GMZ2-specific antibody response increased after the vaccination, but was not correlated to protection. Antibody responses to several P. falciparum blood and liver stage antigens (MSP1, MSP4, MSP8, PfEMP1, STARP) as well as the breadth of the malaria-specific antibody response were significantly higher in protected study participants. CONCLUSIONS: In lifelong malaria exposed adults, the main marker of protection against CHMI is a broad antibody pattern recognizing multiple stages of the plasmodial life cycle. Despite vaccination with GMZ2 using a novel formulation, expansion of the GMZ2-stimulated T cells or the GMZ2-specific B cell response was limited, and the vaccine response could not be identified as a marker of protection against malaria. Trial registration PACTR; PACTR201503001038304; Registered 17 February 2015; https://pactr.samrc.ac.za/TrialDisplay.aspx?TrialID=1038.

Association of low birth weight and polyparasitic infection during pregnancy in Lambaréné, Gabon.

OBJECTIVE: To report the prevalence of polyparasitism during pregnancy in the Lambaréné region of Gabon and its association with newborn birth weight. METHOD: Pregnant women in their third trimester were recruited in a prospective study between November 2011 and March 2015. Parasite infection status was assessed microscopically in stool, urine and blood samples. Maternal demographic and obstetrical characteristics and newborns anthropometric data were collected. Multivariable logistic regression was used to assess the association between low birth weight and polyparasitism. RESULTS: 678 of 927 pregnant women were included for analysis with mean age (SD) of 25 (6.8) years. The analysis showed that 69% (468/678) were infected with at least one parasite (Plasmodium spp., Schistosoma spp., soil-transmitted helminths, filarial infections). This comprised of 38% with monoparasitism and 31% polyparasitism. The proportion of newborn babies with a weight below 2500 g (LBW) in our study was 21% (142/678). Compared to pregnant women without infection, women with monoparasitic infection had adjusted Odds Ratio confidence interval 95% CI (aOR [95%CI]) of 1.6 [0.95-2.73], those with two parasites had aOR 95%CI of 2.63 [1.51-4.62], and those with more than two parasites had aOR of 5.08 [2.5-10.38] for delivering a newborn with low birth weight. CONCLUSION: In Lambaréné, an endemic area for multiple parasite infections, there is a high prevalence of polyparasitism in pregnant women. Polyparasitism is associated with low birth weight. Therefore, there is an urgent need for active screening and treatment of parasite infections in pregnant women to assess the potential public health benefit of such interventions.

Ivermectin for causal malaria prophylaxis: a randomised controlled human infection trial.

OBJECTIVE: Ivermectin is safe and widely used for treating helminth infections. It also kills arthropods feeding on treated subjects, including malaria vectors. Thus, ivermectin mass drug administration as an additional tool for malaria control is being evaluated by WHO. As in vitro data, animal experiments and epidemiological observations suggest that ivermectin has a direct effect on the liver stages of the malaria parasite, this study was designed to assess the prophylactic effect of ivermectin on Plasmodium falciparum controlled human malaria infection. METHODS: A total of 4 volunteers were randomised to placebo, and 8 volunteers were randomised to receive ivermectin 0.4 mg/kg, orally, once 2 h before being experimentally infected intravenously with 3200 P. falciparum sporozoites. The primary endpoint was time to parasitaemia detected by positive thick blood smear; RT-qPCR was performed in parallel. RESULTS: All but one volunteer became thick blood smear positive between day 11 and day 12 after infection, and there was no significant effect of ivermectin on parasitaemia. CONCLUSION: Ivermectin - at the dose used - has no clinically relevant activity against the pre-erythrocytic stages of P. falciparum.

OBJECTIF: L'ivermectine est sûr et largement utilisé pour traiter les helminthiases. Il tue également les arthropodes se nourrissant sur les sujets traités, y compris les vecteurs du paludisme. Ainsi, l'administration en masse d'ivermectine en tant qu'outil supplémentaire de lutte contre le paludisme est actuellement évaluée par l'OMS. Comme les données in vitro, les expériences sur animaux et les observations épidémiologiques suggèrent que l'ivermectine a un effet direct sur les stades hépatiques du parasite du paludisme, cette étude a été conçue pour évaluer l'effet prophylactique de l'ivermectine sur l'infection paludéenne humaine par Plasmodium falciparum contrôlée. MÉTHODES: Quatre volontaires ont été randomisés pour un placebo et 8 volontaires ont été randomisés pour recevoir de l'ivermectine à 0,4 mg/kg en une fois par voie orale, 2 heures avant d'être expérimentalement infectés par voie intraveineuse avec 3.200 sporozoïtes de P. falciparum. Le critère d'évaluation principal était le temps à la parasitémie détectée par un frottis sanguin épais positif. Une RT-qPCR a été réalisée en parallèle. RÉSULTATS: Tous les volontaires sauf un sont devenus positifs pour les frottis sanguins épais entre le jour 11 et le jour 12 après l'infection et il n'y avait aucun effet significatif de l'ivermectine sur la parasitémie. CONCLUSION: L'ivermectine - à la dose utilisée - n'a aucune activité cliniquement pertinente contre les stades pré-érythrocytaires de P. falciparum.

Controlled Human Malaria Infection of Healthy Adults With Lifelong Malaria Exposure to Assess Safety, Immunogenicity, and Efficacy of the Asexual Blood Stage Malaria Vaccine Candidate GMZ2.

BACKGROUND: GMZ2 is a recombinant malaria vaccine inducing immune responses against Plasmodium falciparum (Pf) merozoite surface protein-3 and glutamate-rich protein. We used standardized controlled human malaria infection (CHMI) to assess the efficacy of this asexual blood-stage vaccine. METHODS: We vaccinated 50 healthy, adult volunteers with lifelong exposure to Pf 3 times, at 4-week intervals, with 30 or 100 µg GMZ2 formulated in CAF01, a liposome-based adjuvant; 100 µg GMZ2, formulated in Alhydrogel; or a control vaccine (Verorab). Approximately 13 weeks after the last vaccination, 35/50 volunteers underwent CHMI by direct venous inoculation of 3200 Pf sporozoites (Sanaria® PfSPZ Challenge). RESULTS: Adverse events were similarly distributed between GMZ2 and control vaccinees. Baseline-corrected anti-GMZ2 antibody concentrations 4 weeks after the last vaccination were higher in all 3 GMZ2-vaccinated arms, compared to the control group. All GMZ2 formulations induced similar antibody levels. CHMI resulted in 29/34 (85%) volunteers with Pf parasitemia and 15/34 (44%) with malaria (parasitemia and symptoms). The proportion of participants with malaria (2/5 control, 6/10 GMZ2-Alhydrogel, 2/8 30 µg GMZ2-CAF01, and 5/11 100 µg GMZ2-CAF01) and the time it took them to develop malaria were similar in all groups. Baseline, vaccine-specific antibody concentrations were associated with protection against malaria. CONCLUSIONS: GMZ2 is well tolerated and immunogenic in lifelong-Pf-exposed adults from Gabon, with similar antibody responses regardless of formulation. CHMI showed no protective effect of prior vaccination with GMZ2, although baseline, vaccine-specific antibody concentrations were associated with protection. CHMI with the PfSPZ Challenge is a potent new tool to validate asexual, blood-stage malaria vaccines in Africa. CLINICAL TRIALS REGISTRATION: Pan-African Clinical Trials: PACTR201503001038304.

First-in-human, Randomized, Double-blind Clinical Trial of Differentially Adjuvanted PAMVAC, A Vaccine Candidate to Prevent Pregnancy-associated Malaria.

BACKGROUND: Malaria in pregnancy has major impacts on mother and child health. To complement existing interventions, such as intermittent preventive treatment and use of impregnated bed nets, we developed a malaria vaccine candidate with the aim of reducing sequestration of asexual "blood-stage" parasites in the placenta, the major virulence mechanism. METHODS: The vaccine candidate PAMVAC is based on a recombinant fragment of VAR2CSA, the Plasmodium falciparum protein responsible for binding to the placenta via chondroitin sulfate A (CSA). Healthy, adult malaria-naive volunteers were immunized with 3 intramuscular injections of 20 µg (n = 9) or 50 µg (n = 27) PAMVAC, adjuvanted with Alhydrogel or glucopyranosyl lipid adjuvant in stable emulsion (GLA-SE) or in a liposomal formulation with QS21 (GLA-LSQ). Allocation was random and double blind. The vaccine was given every 4 weeks. Volunteers were observed for 6 months following last immunization. RESULTS: All PAMVAC formulations were safe and well tolerated. A total of 262 adverse events (AEs) occurred, 94 (10 grade 2 and 2 grade 3) at least possibly related to the vaccine. No serious AEs occurred. Distribution and severity of AEs were similar in all arms. PAMVAC was immunogenic in all participants. PAMVAC-specific antibody levels were highest with PAMVAC-GLA-SE. The antibodies inhibited binding of VAR2CSA expressing P. falciparum-infected erythrocytes to CSA in a standardized functional assay. CONCLUSIONS: PAMVAC formulated with Alhydrogel or GLA-based adjuvants was safe, well tolerated, and induced functionally active antibodies. Next, PAMVAC will be assessed in women before first pregnancies in an endemic area. CLINICAL TRIALS REGISTRATION: EudraCT 2015-001827-21; ClinicalTrials.gov NCT02647489.

Immune Responses to a Recombinant Glycoprotein E Herpes Zoster Vaccine in Adults Aged 50 Years or Older.

Background: The herpes zoster subunit vaccine (HZ/su), consisting of varicella-zoster virus glycoprotein E (gE) and AS01B Adjuvant System, was highly efficacious in preventing herpes zoster in the ZOE-50 and ZOE-70 trials. We present immunogenicity results from those trials. Methods: Participants (ZOE-50: ≥50; ZOE-70: ≥70 years of age) received 2 doses of HZ/su or placebo, 2 months apart. Serum anti-gE antibodies and CD4 T cells expressing ≥2 of 4 activation markers assessed (CD42+) after stimulation with gE-peptides were measured in subcohorts for humoral (n = 3293) and cell-mediated (n = 466) immunogenicity. Results: After vaccination, 97.8% of HZ/su and 2.0% of placebo recipients showed a humoral response. Geometric mean anti-gE antibody concentrations increased 39.1-fold and 8.3-fold over baseline in HZ/su recipients at 1 and 36 months post-dose 2, respectively. A gE-specific CD42+ T-cell response was shown in 93.3% of HZ/su and 0% of placebo recipients. Median CD42+ T-cell frequencies increased 24.6-fold (1 month) and 7.9-fold (36 months) over baseline in HZ/su recipients and remained ≥5.6-fold above baseline in all age groups at 36 months. The proportion of CD4 T cells expressing all 4 activation markers increased over time in all age groups. Conclusions: Most HZ/su recipients developed robust immune responses persisting for 3 years following vaccination. Clinical Trials Registration: NCT01165177; NCT01165229.

Mosquito Passage Dramatically Changes var Gene Expression in Controlled Human Plasmodium falciparum Infections.

Virulence of the most deadly malaria parasite Plasmodium falciparum is linked to the variant surface antigen PfEMP1, which is encoded by about 60 var genes per parasite genome. Although the expression of particular variants has been associated with different clinical outcomes, little is known about var gene expression at the onset of infection. By analyzing controlled human malaria infections via quantitative real-time PCR, we show that parasite populations from 18 volunteers expressed virtually identical transcript patterns that were dominated by the subtelomeric var gene group B and, to a lesser extent, group A. Furthermore, major changes in composition and frequency of var gene transcripts were detected between the parental parasite culture that was used to infect mosquitoes and Plasmodia recovered from infected volunteers, suggesting that P. falciparum resets its var gene expression during mosquito passage and starts with the broad expression of a specific subset of var genes when entering the human blood phase.

Recognition of Plasmodium falciparum mature gametocyte-infected erythrocytes by antibodies of semi-immune adults and malaria-exposed children from Gabon.

BACKGROUND: Transmission of malaria from man to mosquito depends on the presence of gametocytes, the sexual stage of Plasmodium parasites in the infected host. Naturally acquired antibodies against gametocytes exist and may play a role in controlling transmission by limiting the gametocyte development in the circulation or by interrupting gamete development and fertilization in the mosquito following ingestion. So far, most studies on antibody responses to sexual stage antigens have focused on a subset of gametocyte-surface antigens, even though inhibitory Ab responses to other gametocyte antigens might also play a role in controlling gametocyte density and fertility. Limited information is available on natural antibody response to the surfaces of gametocyte-infected erythrocytes. METHODS: Ab responses to surface antigens of erythrocytes infected by in vitro differentiated Plasmodium falciparum mature gametocytes were investigated in sera of semi-immune adults and malaria-exposed children. In addition, the effect of immunization with GMZ2, a blood stage malaria vaccine candidate, and the effect of intestinal helminth infection on the development of immunity to gametocytes of P. falciparum was evaluated in malaria-exposed children and adults from Gabon. Serum samples from two Phase I clinical trials conducted in Gabon were analysed by microscopic and flow-cytometric immunofluorescence assay. RESULTS: Adults had a higher Ab response compared to children. Ab reactivity was significantly higher after fixation and permeabilization of parasitized erythrocytes. Following vaccination with the malaria vaccine candidate GMZ2, anti-gametocyte Ab concentration decreased in adults compared to baseline. Ab response to whole asexual stage antigens had a significant but weak positive correlation to anti-gametocyte Ab responses in adults, but not in children. Children infected with Ascaris lumbricoides had a significantly higher anti-gametocyte Ab response compared to non-infected children. CONCLUSION: The current data suggest that antigens exposed on the gametocyte-infected red blood cells are recognized by serum antibodies from malaria-exposed children and semi-immune adults. This anti-gametocyte immune response may be influenced by natural exposure and vaccination. Modulation of the natural immune response to gametocytes by co-infecting parasites should be investigated further and may have an important impact on malaria control strategies.

Maternal mortality cases from pulmonary embolism: A nation-wide study in Turkey.

The aim of the study was to evaluate the maternal mortality cases attributed to pulmonary embolism (PE). PE constituted 7.58% of maternal deaths in 2013. Risk factors for PE were present in 15 (88.2%) of the women. Five women (29.4%) were overweight, and 5 (29.4%) were obese. Four women (23.5%) had cardiac diseases. PE occurred in the postpartum period after caesarean delivery in 9 (52.9%) patients. Eleven (64.7%) of the maternal deaths were recognised as preventable. More deaths attributed to PE occurred in the postpartum period (n = 11) than the antepartum period (n = 5). One other maternal mortality case was after therapeutic abortion. Caesarean section, obesity and cardiac diseases were important risk factors. It can be suggested that monitoring all risk factors and timely recognition of related symptoms and signs with initiation of appropriate management have paramount importance for reducing maternal mortality rate related to pulmonary embolism. Increasing awareness of healthcare professionals as well as the public, and continuously reviewing the cases are also important tools for achieving this goal.

Sporozoite Route of Infection Influences In Vitro var Gene Transcription of Plasmodium falciparum Parasites From Controlled Human Infections.

BACKGROUND: Antigenic variation in Plasmodium falciparum is mediated by the multicopy var gene family. Each parasite possesses about 60 var genes, and switching between active var loci results in antigenic variation. In the current study, the effect of mosquito and host passage on in vitro var gene transcription was investigated. METHODS: Thirty malaria-naive individuals were inoculated by intradermal or intravenous injection with cryopreserved, isogenic NF54 P. falciparum sporozoites (PfSPZ) generated from 1 premosquito culture. Microscopic parasitemia developed in 22 individuals, and 21 in vitro cultures were established. The var gene transcript levels were determined in early and late postpatient cultures and in the premosquito culture. RESULTS: At the early time point, all cultures preferentially transcribed 8 subtelomeric var genes. Intradermal infections had higher var gene transcript levels than intravenous infections and a significantly longer intrahost replication time (P = .03). At the late time point, 9 subtelomeric and 8 central var genes were transcribed at the same levels in almost all cultures. Premosquito and late postpatient cultures transcribed the same subtelomeric and central var genes, except for var2csa CONCLUSIONS: The duration of intrahost replication influences in vitro var gene transcript patterns. Differences between premosquito and postpatient cultures decrease with prolonged in vitro growth.

Impact of adjuvants on CD4(+) T cell and B cell responses to a protein antigen vaccine: Results from a phase II, randomized, multicenter trial.

Immunogenicity and safety of different adjuvants combined with a model antigen (HBsAg) were compared. Healthy HBV-naïve adults were randomized to receive HBs adjuvanted with alum or Adjuvant Systems AS01B, AS01E, AS03A or AS04 at Days 0 and 30. Different frequencies of HBs-specific CD4+ T cells 14days post dose 2 but similar polyfunctionality profiles were induced by the different adjuvants with frequencies significantly higher in the AS01B and AS01E groups than in the other groups. Antibody concentrations 30days post-dose 2 were significantly higher in AS01B, AS01E and AS03A than in other groups. Limited correlations were observed between HBs-specific CD4+ T cell and antibody responses. Injection site pain was the most common solicited local symptom and was more frequent in AS groups than in alum group. Different adjuvants formulated with the same antigen induced different adaptive immune responses and reactogenicity patterns in healthy naïve adults. The results summary for this study (GSK study number 112115 - NCT# NCT00805389) is available on the GSK Clinical Study Register and can be accessed at www.gsk-clinicalstudyregister.com.

Clinical, environmental, and serologic surveillance studies of melioidosis in Gabon, 2012-2013.

Burkholderia pseudomallei, an environmental gram-negative bacillus, is the causative agent of melioidosis and a bio-threat agent. Reports of B. pseudomallei isolation from soil and animals in East and West Africa suggest that melioidosis might be more widely distributed than previously thought. Because it has been found in equatorial areas with tropical climates, we hypothesized that B. pseudomallei could exist in Gabon. During 2012-2013, we conducted a seroprevalance study in which we set up microbiology facilities at a large clinical referral center and prospectively screened all febrile patients by conducting blood cultures and testing for B. pseudomallei and related species; we also determined whether B. pseudomallei could be isolated from soil. We discovered a novel B. pseudomallei sequence type that caused lethal septic shock and identified B. pseudomallei and B. thailandensis in the environment. Our data suggest that melioidosis is emerging in Central Africa but is unrecognized because of the lack of diagnostic microbiology facilities.

Direct venous inoculation of Plasmodium falciparum sporozoites for controlled human malaria infection: a dose-finding trial in two centres.

BACKGROUND: Controlled human malaria infection (CHMI) accelerates development of anti-malarial interventions. So far, CHMI is done by exposure of volunteers to bites of five mosquitoes carrying Plasmodium falciparum sporozoites (PfSPZ), a technique available in only a few centres worldwide. Mosquito-mediated CHMI is logistically complex, exact PfSPZ dosage is impossible and live mosquito-based interventions are not suitable for further clinical development. METHODS: An open-labelled, randomized, dose-finding study in 18-45 year old, healthy, malaria-naïve volunteers was performed to assess if intravenous (IV) injection of 50 to 3,200 aseptic, purified, cryopreserved PfSPZ is safe and achieves infection kinetics comparable to published data of mosquito-mediated CHMI. An independent study site verified the fully infectious dose using direct venous inoculation of PfSPZ. Parasite kinetics were assessed by thick blood smear microscopy and quantitative real time PCR. RESULTS: IV inoculation with 50, 200, 800, or 3,200 PfSPZ led to parasitaemia in 1/3, 1/3, 7/9, and 9/9 volunteers, respectively. The geometric mean pre-patent period (GMPPP) was 11.2 days (range 10.5-12.5) in the 3,200 PfSPZ IV group. Subsequently, six volunteers received 3,200 PfSPZ by direct venous inoculation at an independent investigational site. All six developed parasitaemia (GMPPP: 11.4 days, range: 10.4-12.3). Inoculation of PfSPZ was safe. Infection rate and pre-patent period depended on dose, and injection of 3,200 PfSPZ led to a GMPPP similar to CHMI with five PfSPZ-infected mosquitoes. The infectious dose of PfSPZ predicted dosage of radiation-attenuated PfSPZ required for successful vaccination. CONCLUSIONS: IV inoculation of PfSPZ is safe, well tolerated and highly reproducible. It shall further accelerate development of anti-malarial interventions through standardization and facilitation of CHMI. Beyond this, rational dose selection for whole PfSPZ-based immunization and complex study designs are now possible. TRIAL REGISTRATION: ClinicalTrials.gov NCT01624961 and NCT01771848 .

Respiratory syncytial virus and other respiratory viral infections in older adults with moderate to severe influenza-like illness.

BACKGROUND: Few studies have prospectively assessed viral etiologies of acute respiratory infections in community-based elderly individuals. We assessed viral respiratory pathogens in individuals ≥65 years with influenza-like illness (ILI). METHODS: Multiplex reverse-transcriptase polymerase chain reaction identified viral pathogens in nasal/throat swabs from 556 episodes of moderate-to-severe ILI, defined as ILI with pneumonia, hospitalization, or maximum daily influenza symptom severity score (ISS) >2. Cases were selected from a randomized trial of an adjuvanted vs nonadjuvanted influenza vaccine conducted in elderly adults from 15 countries. RESULTS: Respiratory syncytial virus (RSV) was detected in 7.4% (41/556) moderate-to-severe ILI episodes in elderly adults. Most (39/41) were single infections. There was a significant association between country and RSV detection (P = .004). RSV prevalence was 7.1% (2/28) in ILI with pneumonia, 12.5% (8/64) in ILI with hospitalization, and 6.7% (32/480) in ILI with maximum ISS > 2. Any virus was detected in 320/556 (57.6%) ILI episodes: influenza A (104/556, 18.7%), rhinovirus/enterovirus (82/556, 14.7%), coronavirus and human metapneumovirus (each 32/556, 5.6%). CONCLUSIONS: This first global study providing data on RSV disease in ≥65 year-olds confirms that RSV is an important respiratory pathogen in the elderly. Preventative measures such as vaccination could decrease severe respiratory illnesses and complications in the elderly.

The malaria vaccine candidate GMZ2 elicits functional antibodies in individuals from malaria endemic and non-endemic areas.

BACKGROUND: GMZ2 is a hybrid protein consisting of the N-terminal region of the glutamate-rich protein fused in frame to the C-terminal region of merozoite surface protein 3 (MSP3). GMZ2 formulated in Al(OH)3 has been tested in 3 published phase 1 clinical trials. The GMZ2/alum formulation showed good safety, tolerability, and immunogenicity, but whether antibodies elicited by vaccination are functional is not known. METHODS: Serum samples prior to vaccination and 4 weeks after the last vaccination from the 3 clinical trials were used to perform a comparative assessment of biological activity against Plasmodium falciparum. RESULTS: We showed that the maximum level of immunoglobulin G (IgG) antibodies obtained by GMZ2 vaccination is independent of ethnicity, time under malaria-exposure, and vaccine dose and that GMZ2 elicits high levels of functionally active IgG antibodies. Both, malaria-naive adults and malaria-exposed preschool children elicit vaccine-specific antibodies with broad inhibitory activity against geographically diverse P. falciparum isolates. Peptide-mapping studies of IgG subclass responses identified IgG3 against a peptide derived from MSP3 as the strongest predictor of antibody-dependent cellular inhibition. CONCLUSIONS: These findings suggest that GMZ2 adjuvanted in Al(OH)3 elicits high levels of specific and functional antibodies with the capacity to control parasite multiplication.

Immune responses associated with protection induced by chemoattenuated PfSPZ vaccine in malaria-naive Europeans.

Vaccination of malaria-naive volunteers with a high dose of Plasmodium falciparum sporozoites chemoattenuated by chloroquine (CQ) (PfSPZ-CVac [CQ]) has previously demonstrated full protection against controlled human malaria infection (CHMI). However, lower doses of PfSPZ-CVac [CQ] resulted in incomplete protection. This provides the opportunity to understand the immune mechanisms needed for better vaccine-induced protection by comparing individuals who were protected with those not protected. Using mass cytometry, we characterized immune cell composition and responses of malaria-naive European volunteers who received either lower doses of PfSPZ-CVac [CQ], resulting in 50% protection irrespective of the dose, or a placebo vaccination, with everyone becoming infected following CHMI. Clusters of CD4+ and γδ T cells associated with protection were identified, consistent with their known role in malaria immunity. Additionally, EMRA CD8+ T cells and CD56+CD8+ T cell clusters were associated with protection. In a cohort from a malaria-endemic area in Gabon, these CD8+ T cell clusters were also associated with parasitemia control in individuals with lifelong exposure to malaria. Upon stimulation with P. falciparum-infected erythrocytes, CD4+, γδ, and EMRA CD8+ T cells produced IFN-γ and/or TNF, indicating their ability to mediate responses that eliminate malaria parasites.

Safety and efficacy of dihydroartemisinin-piperaquine for intermittent preventive treatment of malaria in pregnant women with HIV from Gabon and Mozambique: a randomised, double-blind, placebo-controlled trial.

BACKGROUND: The cornerstone of malaria prevention in pregnancy, intermittent preventive treatment (IPTp) with sulfadoxine-pyrimethamine, is contraindicated in women with HIV who are receiving co-trimoxazole prophylaxis. We assessed whether IPTp with dihydroartemisinin-piperaquine is safe and effective in reducing the risk of malaria infection in women with HIV receiving co-trimoxazole prophylaxis and antiretroviral drugs. METHODS: For this randomised, double-blind, placebo-controlled clinical trial, women with HIV attending the first antenatal care clinic visit, resident in the study area, and with a gestational age up to 28 weeks were enrolled at five sites in Gabon and Mozambique. Participants were randomly assigned (1:1) to receive either IPTp with dihydroartemisinin-piperaquine at each scheduled antenatal care visit plus daily co-trimoxazole (intervention group) or placebo at each scheduled antenatal care visit plus daily co-trimoxazole (control group). Randomisation was done centrally via block randomisation (block sizes of eight), stratified by country. IPTp was given over 3 days under direct observation by masked study personnel. The number of daily IPTp tablets was based on bodyweight and according to the treatment guidelines set by WHO (target dose of 4 mg/kg per day [range 2-10 mg/kg per day] of dihydroartemisinin and 18 mg/kg per day [range 16-27 mg/kg per day] of piperaquine given once a day for 3 days). At enrolment, all participants received co-trimoxazole (fixed combination drug containing 800 mg trimethoprim and 160 mg sulfamethoxazole) for daily intake. The primary study outcome was prevalence of peripheral parasitaemia detected by microscopy at delivery. The modified intention-to-treat population included all randomly assigned women who had data for the primary outcome. Secondary outcomes included frequency of adverse events, incidence of clinical malaria during pregnancy, and frequency of poor pregnancy outcomes. All study personnel, investigators, outcome assessors, data analysts, and participants were masked to treatment assignment. This study is registered with ClinicalTrials.gov, NCT03671109. FINDINGS: From Sept 18, 2019, to Nov 26, 2021, 666 women (mean age 28·5 years [SD 6·4]) were enrolled and randomly assigned to the intervention (n=332) and control (n=334) groups. 294 women in the intervention group and 308 women in the control group had peripheral blood samples taken at delivery and were included in the primary analysis. Peripheral parasitaemia at delivery was detected in one (<1%) of 294 women in the intervention group and none of 308 women in the control group. The incidence of clinical malaria during pregnancy was lower in the intervention group than in the control group (one episode in the intervention group vs six in the control group; relative risk [RR] 0·12, 95% CI 0·03-0·52, p=0·045). In a post-hoc analysis, the composite outcome of overall malaria infection (detected by any diagnostic test during pregnancy or delivery) was lower in the intervention group than in the control group (14 [5%] of 311 women vs 31 [10%] of 320 women; RR 0·48, 95% CI 0·27-0·84, p=0·010). The frequency of serious adverse events and poor pregnancy outcomes (such as miscarriages, stillbirths, premature births, and congenital malformations) did not differ between groups. The most frequently reported drug-related adverse events were gastrointestinal disorder (reported in less than 4% of participants) and headache (reported in less than 2% of participants), with no differences between study groups. INTERPRETATION: In the context of low malaria transmission, the addition of IPTp with dihydroartemisinin-piperaquine to co-trimoxazole prophylaxis in pregnant women with HIV did not reduce peripheral parasitaemia at delivery. However, the intervention was safe and associated with a decreased risk of clinical malaria and overall Plasmodium falciparum infection, so it should be considered as a strategy to protect pregnant women with HIV from malaria. FUNDING: European and Developing Countries Clinical Trials Partnership 2 (EDCTP2) and Medicines for Malaria Venture. TRANSLATIONS: For the Portuguese and French translations of the abstract see Supplementary Materials section.